Regulation of parasite-specific immune responses in filarial infection- Using a variety of techniques including cell culture, ELISPOT, RT-PCR, and flow cytometry-based intracellular detection of cytokines, we were able to demonstrate that the ?defect? in the immunological responsiveness to parasite antigen in the asymptomatic microfilarial-positive individuals was a profound inability to produce cytokines associated with a Type-1 response, IL-2 and IFN-g most notably. Further, it was shown that IL-10 was the cytokine responsible for the parasite-specific down- regulation seen. Although Type-2 T cells can make IL-10, another important source of the IL-10 in these individuals appears to the nonlymphocyte cell populations (particularly the monocytes and other antigen-presenting cells) based on IL-10 protein measurement and mRNA production in immunomagnetically separated cell populations. The ability to overcome the Ag-specific hyporesponsiveness was also studied in vitro by assessing the ability of neutralizing anti-IL-10 antibody or recombinant IL-12 to restore the production of Type1-type cytokines in hyporesponsive individuals. Both anti-IL-10 and rIL-12 were able to augment the production of IFN-g in response to parasite Ag. These studies indicated that IL-10 plays a major role in the maintenance of Ag-specific hyporesponsiveness in microfilaremic individuals, and IL-12 may play a significant role in overcoming this hyporesponsiveness. Furthermore, using stage-specific antigenic preparation, the hyporesponsiveness and the induction of IL-10 appears to be preferentially induced by microfilarial antigens; this microfilarial antigen-induced IL-10 appears to work by diminishing the expression of the important costimulatory molecules, CD80/86 and its counter ligand CD28. Because chronic filarial infections may alter immune reactivity to other (nonparasite) antigens and because these alterations may have profound implications for vaccine programs worldwide, a community based study in Esmeraldas province of Ecuador was performed in which 200 individuals (mostly children) with microfilaria-positive onchocerciasis along with ethnically similar but uninfected individuals were vaccinated with common, commercially available vaccines (tetanus toxoid as a ?recall? protein antigen, pneumococcal vaccine as a new and/or a recall polysaccharide, hepatitis B as a new protein), and their subsequent cellular (cytokine profile) and humoral (isotype, subclass) responses were compared. Preliminary examination of the cellular response demonstrates very clearly that the presence of active onchocerciasis blunts significantly the proliferative response along with IFN-gamma production to tetanus toxoid and changes the isotype profile of the antibody responses postvaccination to hepatitis B and to tetanus. Predisposition to Infection and Disease - To examine the possibility that there is a genetic predisposition to a given immune response or to a given pathological outcome of infection, novel methods of examining the MHC class II genetic diversity have been developed and utilized in the fine mapping of the DQ, DP and DR locus in <600 patients with varying forms of filarial infections. To date, almost all of the HLA Class II alleles have been typed in these many individuals. Of note, there is a statistically significant difference between putatively immune (PI) and infected (INF) Amerindian subjects. This finding appears to be the result of a polarization of HLA-DR -DQ haplotypes DRB1*08042-DQA1*0401-DQB1*0402 (PI) and DRB1*0404- DQA1*0301-DQB1*0302 (INF). The presence of an association between HLA class II genes and immunity or infection has not been observed within the South American Black population subset that cohabit this onchocerca-endemic region, however. We have developed several in vitro models to study this early immune response. We have been able to isolate in pure form from normal, unsensitized blood donors both dendritic cells (the first cell likely to encounter a parasite antigen) and monocytes and shown that, upon stimulation with the larval stage antigen, parasite antigen induces these cells to produce IL-10 in significantly greater amounts than is produced to the non- helminth protozoal antigen, soluble Toxoplasma Ag. Further, in response to filarial antigens, these dendritic cells and monocytes produce RANTES, one of the major eosinophil chemoattractant chemokines. Most recently, we have shown that parasite antigen and live parasites preferentially induce IL-4 production by dendritic cells, a finding that has profound implications for understanding why the T cell response in these infections invariably leads to immune responses characterized by IL-4 and IL-5. Modeling the early immune response to parasitic helminth infection - We have also developed a model to examine both priming and induction of an immune response to parasite antigen in CD4+CD45RA+ (naive) T cells obtained from unsensitized donors. We have shown that these ?naive? CD4+ cells were able to proliferate and produce IL-5 and IFN-g when primed with microfilarial antigen using B-cells/monocytes as APCs. This same microfilarial Ag in the presence of IL-4 induces a strong IL-5 (Type-2) response whereas in its absence a Th0 (both IL-5 and IFN-g) occurs. On secondary stimulation with microfilarial antigen, cells originally primed with Mf Ag or Mf Ag + IL-2 produced IFN-g whereas cells primed in the presence of IL-4, a Th2-type response was induced.